MicroPET, PET for small animals, has recently become an important in vivo imaging technique for studying biology and for drug evaluation/development for many medical disorders, including cancer. The number of microPET scanners in operation is growing rapidly. However, the full potential of microPET cannot be realized without the ability to provide absolute quantification of biological function that usually requires the time activity curve of tracer in blood/plasma (input function), which is difficult to obtain in small animals like the mouse. The goal of this study is thus to establish and validate image-based methods that can provide the input function in these small animals, that in turn allows absolute biological quantitation with microPET studies. In this application, a set of microPET studies will be performed with four commonly used microPET tracers (i.e., FDG, FLT, FHBG, and N-13 ammonia) to provide data that are usable for validation of image-based input function derivation methods. A novel validation method that does not require many serial blood samples to be taken from a single animal will be used for evaluation and validation of various input function derivation methods (including factor analysis, whole body modeling, standard shape, and blind de-convolution). Based on the microPET data collected in this study, investigation will also be performed to search for surrogate input functions for these common tracers to develop simple and practical quantitation procedures. Results from the proposed study will enable microPET to provide quantitative and reliable biological functions in local tissue regions in mouse microPET studies that are essential for understanding and treatment of many disorders, including cancer and cardiovascular diseases, and for drug development. The full set of image data from microPET experiments will also be made available to the public on the internet for other investigators to evaluate and validate their new/favorable input function derivation methods.